This application is a 371 of PCT/GB96/02134 filed Aug. 28, 1996.
This invention relates to the interconversion of inactive 11-keto steroids with their active 11xcex2-hydroxy equivalents, to methods by which the conversion of the inactive to the active form may be controlled, and to useful therapeutic effects which may be obtained as a result of such control. More specifically, but not exclusively, the invention is concerned with interconversion between cortisone and cortisol in humans.
Glucocorticoids such as cortisol have a number of diverse effects on different body tissues. For example, the use of cortisol as an anti-inflammatory agent was described in our International Patent Application WO 90/04399, which was concerned with the problem that therapeutically administered cortisol tends to be converted in the body to inactive cortisone by 11xcex2-hydroxysteroid dehydrogenase enzymes. Our earlier invention provided for the potentiation of cortisol by the administration of an inhibitor of the 11xcex2-dehydrogenase activity of these enzymes.
Another major physiological effect of cortisol is its antagonism to insulin, and it is known for example that high concentrations of cortisol in the liver substantially reduce insulin sensitivity in that organ, thus tending to increase gluconeogenesis and consequently raising blood sugar levels [1]. This effect is particularly disadvantageous in patients suffering from impaired glucose tolerance or diabetes mellitus, in whom the action of cortisol can serve to exacerbate insulin resistance. Indeed, in Cushing""s syndrome, which is caused by excessive circulating concentrations of cortisol, the antagonism of insulin can provoke diabetes mellitus in susceptible individuals [2].
As mentioned above, it is known that cortisol can be converted in the body to cortisone by the 11xcex2-dehydrogenase activity of 11xcex2-hydroxysteroid dehydrogenase enzymes. It is also known that the reverse reaction, converting inactive cortisone to active cortisol, is accomplished in certain organs by 11xcex2-reductase activity of these enzymes. This activity is also known as corticosteroid 11xcex2-reductase, cortisone 11xcex2-reductase, or corticosteroid 11xcex2-oxidoreductase.
It has only recently become apparent that there are at least two distinct isozymes of 11xcex2-hydroxysteroid dehydrogenase (collectively abbreviated as 11xcex2-HSD, which term is used, where appropriate, in this specification). Aldosterone target organs and placenta express a high affinity NAD+-dependent enzyme (11xcex2-HSD2) [3]. This has been characterised in placenta and kidney [4,5] and cDNA clones have been isolated [6,9]. 11xcex2-HSD2 catalyses 11xcex2-dehydrogenase activity exclusively [4,7]. In contrast, the previously purified, liver derived isozyme (11xcex2-HSD1) is a lower affinity, NADP+/NADPH-dependent enzyme [10,11]. Expression of 11xcex2-HSD1 in a range of cell lines encodes either a bi-directional enzyme [11,12] or a predominant 11xcex2-reductase [13,15] which, far from inactivating glucocorticoids, regenerates active 11xcex2-hydroxysteroid from otherwise inert 11-keto steroid. 11xcex2-reductase activity, best observed in intact cells, activates 11-keto steroid to alter target gene transcription and differentiated cell function [13,14]. 11xcex2-HSD1 and 11xcex2-HSD2 are the products of different genes and share only 20% amino acid homology [6,7].
As far as the applicants are aware no previous attempts have been made to modify the action of 11xcex2-reductase. We have now found that it is possible to inhibit this activity in vivo, and in doing so we have created the possibility of a novel medicament for use in treating many of the deleterious effects of glucocorticoid excess. In one aspect, therefore, the invention provides the use of an inhibitor of 11xcex2-reductase in the manufacture of a medicament for control of 11-keto steroid conversion to 11xcex2-hydroxysteroid in vivo.
As mentioned above, one of the major physiological effects of cortisol is insulin antagonism in the liver, and in a specific aspect the invention therefore provides the use of an inhibitor of 11xcex2-reductase in the manufacture of a medicament for inhibiting hepatic gluconeogenesis [1]. Cortisol promotes hepatic gluconeogenesis by several mechanisms, including antagonism of the effects of insulin on glucose transport, and interactions with insulin and glucose in the regulation of several enzymes which control glycolysis and gluconeogenesis. These include glucokinase, 6-phosphofructokinase, pyruvate kinase, phosphoenolpyruvate carboxykinase (PEPCK), and glucose-6-phosphatase. Inhibiting production of cortisol from cortisone in the liver therefore enhances hepatic glucose uptake and inhibits hepatic glucose production by several mechanisms [16]. Moreover, the influence of inhibiting 11xcex2-reductase activity in the liver of patients with insulin resistance or glucose intolerance may be greater than in healthy subjects because in insulin resistance or deficiency the influence of cortisol on PEPCK has been shown to be greater [17]; obese patients secrete more cortisol [18]; insulin resistant patients are more sensitive to glucocorticoids [19]; and insulin down-regulates 11xcex2-HSD1 expression [15,20] so that 11xcex2-reductase activity may be enhanced in conditions of insulin resistance or deficiency.
Our studies have also shown that 11xcex2-HSD1 is expressed in rat adipose tissue and in adipocyte cell lines in culture, where it converts 11-dehydrocorticosterone to corticosterone (the rat equivalents of human cortisone and cortisol, respectively). This suggests that similar 11xcex2-reductase activity will be observed in human adipose tissue, with the result that inhibition of the enzyme will result in alleviation of the effects of insulin resistance in adipose tissue in humans. This would lead to greater tissue utilisation of glucose and fatty acids, thus reducing circulating levels. The invention therefore provides, in a further aspect, the use of an inhibitor of 11xcex2-reductase in the manufacture of a medicament for increasing insulin sensitivity in adipose tissue.
The results of our studies have encouraged us to believe that inhibition of intracellular cortisol production will also lead to increased insulin sensitivity in other tissues which are acted upon by insulin, for instance skeletal muscle [21]. Inhibiting the 11xcex2-reductase therefore promises to reverse the effects of insulin resistance in muscle tissue, and to promote the up-take of essential molecules such as glucose and free fatty acids into muscle cells with consequent improved muscle metabolism and reduction of circulating levels of glucose and fatty acids. In a further aspect, the invention therefore provides the use of an inhibitor of 11xcex2-reductase in the manufacture of a medicament for increasing insulin sensitivity in skeletal muscle tissue.
It is also known that glucocorticoid excess potentiates the action of certain neurotoxins, which leads to neuronal dysfunction and loss. We have studied the interconversion between 11-dehydrocorticosterone and corticosterone in rat hippocampal cultures, and have found (surprisingly in view of the damaging effects of glucocorticoids) that 11xcex2-reductase activity dominates over 11xcex2-dehydrogenase activity in intact hippocampal cells [22]. The reason for this activity is unknown, but this result indicates that glucocorticoid excess may be controlled in hippocampal cells (and by extension in the nervous system in general) by use of an 11xcex2-reductase inhibitor, and the invention therefore provides in an alternative aspect the use of an inhibitor of 11xcex2-reductase in the manufacture of a medicament for the prevention or reduction of neuronal dysfunction and loss due to glucocorticoid potentiated neurotoxicity. It is also possible that glucocorticoids are involved in the cognitive impairment of ageing with or without neuronal loss and also in dendritic attenuation [23-25]. Furthermore, glucocorticoids have been implicated in the neuroual dysfunction of major depression. Thus an inhibitor of 11xcex2-reductase could also be of value in these conditions.
It will be appreciated from the foregoing that the potential beneficial effects of inhibitors of 11xcex2-reductase are many and diverse, and it is envisaged that in many cases a combined activity will be demonstrated, tending to relieve the effects of endogenous glucocorticoids in diabetes mellitus, obesity (including centripetal obesity), neuronal loss and the cognitive impairment of old age. Thus, in a further aspect, the invention provides the use of an inhibitor of 11xcex2-reductase in the manufacture of a medicament for producing multiple therapeutic effects in a patient to whom the medicament is administered, said therapeutic effects including an inhibition of hepatic gluconeogenesis, an increase in insulin sensitivity in adipose tissue and muscle, and the prevention of or reduction in neuronal loss/cognitive impairment due to glucocorticoid-potentiated neurotoxicity or neural dysfunction or damage.
From an alternative point of view, the invention provides a method of treatment of a human or animal patient suffering from a condition selected from the group consisting of: hepatic insulin resistance, adipose tissue insulin resistance, muscle insulin resistance, neuronal loss or dysfunction due to glucocorticoid potentiated neurotoxicity, and any combination of the aforementioned conditions, the method comprising the step of administering to said patient a medicament comprising a pharmaceutically active amount of an inhibitor of 11xcex2-reductase.
As mentioned previously, the factors which control the relative activities of 11xcex2-dehydrogenase and 11xcex2-reductase in different conditions, especially by the 11xcex2-HSD1 isozyme, are poorly understood. It is likely that an 11xcex2-reductase inhibitor will be selective for the 11xcex2-HSD1 isozyme in vivo. We have found, for instance, that glycyrrhetinic acid (a known inhibitor of 11xcex2-dehydrogenase) has no effect on 11xcex2-reductase in vivo [26]. However, we have surprisingly found that carbenoxolone, which is known as an inhibitor of the 11xcex2-dehydrogenase enzyme, also inhibits 11xcex2-reductase in vivo [26,27]. In preferred embodiments, therefore, the inhibitor is carbenoxolone (3xcex2-(3-carboxypropionyloxy)-11-oxo-olean-2-en 30-oic acid), or a pharmaceutically acceptable salt thereof. The dose of carbenoxolone which we used in our studies was 100 mg every 8 hours given orally.